Conditional expression of the mitogen-activated protein kinase (MAPK) phosphatase MKP-1 preferentially inhibits p38 MAPK and stress-activated protein kinase in U937 cells
Conditional expression of the mitogen-activated protein kinase (MAPK) phosphatase MKP-1 preferentially inhibits p38 MAPK and stress-activated protein kinase in U937 cells
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DOI：
10.1074/jbc.272.27.16917
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发表时间：
1997-07-04
影响因子：
4.8
通讯作者：
Kraft, AS
中科院分区：
文献类型：
--
作者：
Franklin, CC;Kraft, AS
佛波酯肿瘤促进剂，如佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA），是U937人白血病细胞中细胞外信号调节激酶2（ERK 2）、应激活化蛋白激酶（SAPK）和p38促分裂原活化蛋白激酶（MAPK）的有效激活剂。这些激酶受保守的苏氨酸和酪氨酸残基的可逆双重磷酸化调节，双特异性蛋白磷酸酶MAPK磷酸酶-I（MKP-1）已显示在瞬时转染研究中去磷酸化和活化ERK 2、SAPK和p38 MAPK。在此，我们证明PMA处理诱导U937细胞中的MKP-1蛋白表达，其在30分钟内可检测到，在4小时后达到最大水平。该时间过程与PMA诱导的SAPK活性的快速失活相一致，但与ERK 2磷酸化不一致，ERK 2磷酸化保持升高长达6小时。我们建立了从人金属硫蛋白IIa启动子条件性表达MKP-1的U937细胞系，MKP-1的条件性表达抑制PMA诱导的ERK 2、SAPK和p38 MAPK活性。通过滴定从人金属硫蛋白IIa启动子表达MKP-1的水平，然而，发现p38 MAPK和SAPK对MKP-1的抑制比ERK 2敏感得多，MKP-1的这种不同底物特异性可以在功能上扩展到核转录事件，因为PMA诱导的c-Jun转录活性对MKP-1的抑制比Elk-1或c-Myc更敏感。MKP-1的条件性表达也消除了响应PMA处理的内源性MKP-1蛋白表达的诱导，这种负反馈调节机制可能是由于MKP-1介导的ERK 2抑制，因为利用MEK 1/2抑制剂PD98059的研究表明，ERK 2活化是PMA诱导的MKP-1表达所必需的。这些结果表明，ERK 2介导的诱导MKP-1可能发挥重要作用，优先衰减信号通过p38 MAPK和SAPK信号转导途径。
Phorbol ester tumor promoters, such as phorbol 12-myristate 13-acetate (PMA), are potent activators of extracellular signal-regulated kinase 2 (ERK2), stress-activated protein kinase (SAPK), and p38 mitogen-activated protein kinase (MAPK) in U937 human leukemic cells, These kinases are regulated by the reversible dual phosphorylation of conserved threonine and tyrosine residues, The dual specificity protein phosphatase MAPK phosphatase-l (MKP-1) has been shown to dephosphorylate and inactivate ERK2, SAPK, and p38 MAPK in transient transfection studies. Here we demonstrate that PMA treatment induces MKP-1 protein expression in U937 cells, which is detectable within 30 min with maximal levels attained after 4 h, This time course coincides with the rapid inactivation of PMA-induced SAPK activity, but not ERK2 phosphorylation, which remains elevated for up to 6 h, To examine directly the role of MKP-1 in the regulation of these protein kinases in vivo, we established a U937 cell line that conditionally expresses MKP-1 from the human metallothionein IIa promoter, Conditional expression of MKP-1 inhibited PMA-induced ERK2, SAPK, and p38 MAPK activity, By titrating the levels of MKP-1 expression from the human metallothionein IIa promoter, however, it was found that p38 MAPK and SAPK were much more sensitive to inhibition by MKP-1 than ERK2, This differential substrate specificity of MKP-1 can be functionally extended to nuclear transcriptional events in that PMA-induced c-Jun transcriptional activity was more sensitive to inhibition by MKP-1 than either Elk-l or c-Myc. Conditional expression of MKP-1 also abolished the induction of endogenous MKP-1 protein expression in response to PMA treatment, This negative feedback regulatory mechanism is likely due to MKP-1-mediated inhibition of ERK2, as studies utilizing the MEK1/2 inhibitor PD98059 suggest that ERK2 activation is required for PMA-induced MKP-1 expression, These findings suggest that ERK2-mediated induction of MKP-1 may play an important role in preferentially attenuating signaling through the p38 MAPK and SAPK signal transduction pathways.